Over-expression of ubiquitin carboxy terminal hydrolase-L1 induces apoptosis in breast cancer cells.
Ubiquitin carboxy terminal hydrolase-L1 (UCH-L1) belongs to the UCH proteases family that deubiquitinates ubiquitin-protein conjugates in the ubiquitin-proteasome system. Previous research showed that UCH-L1 was expressed in mouse retinal cells and testicular germ cells, and its function was associated with apoptosis. But it is still unclear whether UCH-L1 is concerned with apoptosis in tumor cells. In order to clarify the role of UCH-L1 in tumor cells, multi-drug resistance (MDR) human breast carcinoma cell line MCF7/Adr, that expresses relatively high UCH-L1, and its parental cell line MCF7, that expresses relatively low UCH-L1, were chosen for this study. We transfected pcDNA3.1-UCH-L1 plasmid and UCH-L1 siRNA into MCF7 and MCF7/Adr cells, respectively. Using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, western blot, Hoechst 33258 staining assay and flow cytometry, we found that over-expression of UCH-L1 in MCF7 cells induced apoptosis. On the other hand, silencing of UCH-L1 in MCF7/Adr cells led to the opposite effect. Moreover, to explore the mechanism underling these observations, we further investigated the expression of phospho-Akt and its downstream signal phospho-IkB-alpha and other signal molecules including Fas, Fas-L, Trail, DR4, DR5, Bax, cytochrome C, active caspase-3, phospho-p53, phospho-Mdm-2, Bcl-2, Bcl-xL, p21 and p27. The results indicated that the process of apoptosis triggered by UCH-L1 is, at least in part, probably through Phosphoinositide 3-kinase (PI3K)/Akt signal pathway. Our findings suggest that modulating the ubiquitination and deubiquitination pathway could be a novel method for tumor therapy.
Wang
Wen-Juan
WJ
Department of Pathology, Shanghai Medical College, Fudan University, Shanghai 200032, PR China.
Li
Qing-Quan
QQ
Xu
Jing-Da
JD
Cao
Xi-Xi
XX
Li
Hai-Xia
HX
Tang
Feng
F
Chen
Qi
Q
Yang
Jin-Ming
JM
Xu
Zu-De
ZD
Liu
Xiu-Ping
XP
D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D050756:Cyclin-Dependent Kinase Inhibitor Proteins / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D005260:Female; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D043222:Ubiquitin Thiolesterase / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation
D051017:Apoptosis Regulatory Proteins; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D020672:I-kappa B Proteins; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D034741:RNA, Small Interfering; C507533:Ubiquitin carboxyl-Terminal Hydrolase L-1, human; D000072000:NF-KappaB Inhibitor alpha; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D043222:Ubiquitin Thiolesterase